GEN 604 EXAM 4

Genetic switches

A molecular system that controls the expression of a specific gene

OR

Places in genome that are regulated by regulatory proteins; same as cis regulatory elements

\______ drives gene expression

Protein-DNA interactions

The binding of \_____ or \_____ to \_______ determines if transcription will take place

activators; repressors; operators

Positive regulation

An activator is involved in recruiting RNA polymerase, gene is transcribed

Negative regulation

A repressor prevents the recruitment of RNA polymerase, gene is not transcribed

DNA-binding domain

Feature in activators/repressors that binds to DNA

Allosteric site

Feature in activators/repressors that bind an environmental signal (allosteric effector/inducer) that leads to a change in the protein's shape.

Allosteric effector

A small molecule that binds to an allosteric site of an activator or repressor, causing it to change shape. Aka inducer

Most loci are under...

Both positive and negative regulation

O site

Operator. Where repressor protein binds

P site

Promoter

Activator and repressor proteins

Sensors that monitor the environment and impact gene expression accordingly

lac structural genes (Z, Y, A)

Encode enzymes for the metabolism of lactose

lacZ

Encodes B-galactosidase

B-galactosidase

Enzyme that breaks lactose down into glucose and galactose OR allolactose

lacY

Encodes lactose permease

Lactose permease

Membrane protein that transports lactose into the cell

lacA

Encodes transacetylase

Operon

A set of adjacent genes whose mRNA is synthesized in one piece, along with any adjacent regulatory sites. Genes are coordinately controlled.

Allosteric transition

A change from one conformation of a protein to another. Occurs when an allosteric effector binds.

Induction

Relief of repression

CAP protein

Key regulatory molecule involved in positive gene regulation of the lac operon. Binds to cAMP

cAMP-CAP complex

Bind to P site to activate gene expression (recruits RNA polymerase)

Rotational symmetry

Based on palindrome sequences. Occurs as a consequence of DNA's double-helical structure

Palindromes are often...

Regulatory sites

ara Structural genes

B, A, D.

C

Control gene

ara Control sites

O, I, P

AraC

Protein that is a dual regulator of the arabinose operon. When bound to arabinose, binds to araI and allows for transcription. When not bound to arabinose, binds to both araI and araO, blocking RNA polymerase from transcription

Trp operon

Five genes for the synthesis of tryptophan

Trp repressor

Represses transcription when tryptophan is present in the environment

Attenuation

Transcriptional control that functions by premature termination of mRNA synthesis. Occurs based on level of translation.

Leader sequence of 5'-UTR

Region of mRNA that accomplishes attenuation. In trp operon, has two tryptophan codons in a row.

Attenuation when tryptophan is present

Ribosome easily passes two tryptophan codons -\> the mRNA being created forms a stem loop structure that leads to the early termination of translation and transcription

Attenuation when tryptophan is absent

Ribosome stalls at the two tryptophan codons -\> mRNA does not form stem loop structure, allowing for the translation of tryptophan-synthesizing molecules

Phage lambda

Bacteriophage that infects E. coli that can have two distinct life cycles

Lytic cycle

Phage production and lysis. Preferred when host cell is in good conditions, experiencing high rates of cell division.

Lysogenic cycle

Lambda genome becomes integrated into the host. Preferred when host cell is in poor conditions, experiencing low rates of cell division.

cI

Encodes λ repressor. Represses the lytic cycle, promotes the lysogenic cycle.

cro

Encodes Cro repressor. Represses the lysogenic cycle, promotes the lytic cycle. Located on PR

Upon infection with phage λ, transcription is initiated...

At PR (promoter right) and PL (promoter left)

PR (promoter right)

Includes genes for lysis of host, genes for creating phage components (head and tail)

PL (promoter left)

Includes genes involved in recombination, integrases and excisionases

N

Antiterminator. Allows all transcription/translation to occur without termination - all λ genes encoded.

cII

Activates transcription of cI. Very sensitive to proteases -\> indirectly senses the host's environment.

Phage λ in good conditions

High levels of protease -\> low levels of cII -\> low levels of cI -\> Cro repressor represses lysogeny

Phage λ in poor conditions

Low levels of protease -\> high levels of cII -\> high levels of cI -\> λ repressor shuts down transcription of PR and PL (Cro not expressed)

The two repressors of phage λ...

Bind at either the left or right operators to direct RNA polymerase one way or another

Helix-turn-helix

Protein domains that are a common DNA-binding motif. Fits into major groove of DNA.

Enhancers

Regulatory DNA sequences that are distinct from promoters. Bound by transcription factors. In eukaryotes.

Transcription factors (TFs)

Regulatory proteins that bind enhancers

General transcription factors (GTFs)

Regulatory proteins that bind to the core promoter (not enhancers!)

Core promoter

DNA sequence that includes the start point of transcription

Coactivators

Serve as a bridge between TFs and RNA polymerase. Do not bind directly to DNA

Corepressors

Block TFs and RNA polymerase from interacting. Do not bind directly to DNA

Cis regulatory elements

Always exist as DNA within the sequence of the gene being regulated. Same as genetic switches

Trans regulatory elements

Any regulatory element that is not part of a gene's DNA sequence. All regulatory proteins are trans regulatory elements.

Negative switches

Are turned on by repressors

Positive switches

Are turned on by activators

Coactivators and corepressors...

Do not directly bind to DNA, but instead bind to DNA-binding molecules

Proximal enhancer elements

Close to promoter, bound by TFs and required for transcription

Domains of transcription factors (TFs)

•DNA-binding domain

•Dimerization domain

•Ligand-binding domain

•Activation/repression domains

Dimerization domain

Allows TFs to dimerize

Ligand-binding domain

Allows TFs to bind to ligands (often other proteins)

Activation/repression domain

When bound, changes the TF's function

Gal pathway

Pathway in yeast involved in the metabolism of galactose. Includes a number of genes involved with transport and glycolysis.

Gal4

Main transcription factor in the Gal pathway. Has multiple functional domains: DNA-binding and activation.

Upstream activation sequence (UAS)

The enhancer that Gal4 binds to.

When the activation domain of Gal4 is removed...

There is no transcription of lacZ (reporter gene)

Reporter gene

Gene encoding a protein whose activity is easy to monitor experimentally; used to study the expression pattern of a target gene or the localization of its protein product.

Gal80

Constitutively expressed and prevents Gal4 activity in the absence of galactose by binding to Gal4. Released from DNA upon binding of galactose-Gal3 complex.

\___ is an allosteric effector of Gal3, and \_____ is an allosteric effector of Gal80.

Galactose; Gal3

Two ways activators work indirectly to initiate transcription

1\.) bind to TBP, allowing TFIID to bind

2\.) recruits a mediator complex (a coactivator) that itself recruits RNA polymerase II

Histones

Proteins that chromatin is composed of

H1

Linker histone. Not part of nucleosomes

H2A, H2B, H3, H4

Core histones. Part of nucleosomes.

Nucleosome

Main unit of chromatin. Made up of four core histones linked together.

Chromatin modification

Chemical modification of chromatin; alter interaction with DNA

Chromatin remodeling

Sliding, ejecting, removing, or replacing histones that comprise nucleosomes. Alter the abilities of TFs to bind.

Histone acetyl transferase (HAT)

Adds acetyl groups to lysine residues in histones.

Histone acetylation...

Loosens DNA wrapping and a spreads out the histones and increases gene expression

Histone methylation...

Tightens DNA wrapping also silences gene expression

Histone code

The pattern of modifications of histones that affects the chromatin structure and gene transcription. The cell can interpret the epigenetic elements that make up the histone code to know what to do.

DNA methyltransferases (DNMTs)

Add methyl groups to DNA

TET enzymes

Remove methyl groups from DNA

Methylation occurs on \_____ residues

Lysine or arginine

The histone code....

Tells the cell where transcription should occur, based on histone modification patterns

DNA methylation

Another way to repress transcription. DNA itself is directly methylated.

Reader proteins are \____ while writers, erasers, and chromatin remodelers are \_____

TFs; enzymes

Heterochromatin

Highly condensed chromatin. Transcriptionally inactive, for genes that are rarely needed

Euchromatin

Loosely packed chromatic. Transcriptionally active, for genes that are likely to be needed.

Epigenetic inheritance

Chromatin states can be inherited.

Epigenetic marks

Guide the modification of new histones after replication

Position effect

Where a gene is can determine expression - if a gene is near or part of heterchromatin, it may not be expressed.

Genomic imprinting

When an allele inherited by one parent is preferentially expressed while the allele from the other parent is silent. An epigenetic mechanism based on DNA methylation

Insulin-like growth factor 2 (IGF2) is always expressed from the \______ allele, while H19 is always expressed from the \_____ allele

father's; mother's

Dosage compensation

Silences the genes on one chromosome. Occurs in females - one X chromosome is silenced.

Organizers

Tissues capable of controlling the development of other tissues. The tissues express a set of genes that allow for the initiation of development.

Questions of developmental genetics

1\.) What genes control development?

2\.) When and where are these genes expressed?

3\.) How are these genes regulated?

4\.) What is the function of the gene product(s)?

Housekeeping genes

Control metabolism, biosynthesis, etc. Gene products involved in homeostasis.